Several studies clearly demonstrate that the rat submandibular gland is responsive to insulin stimulation both in vivo and in vitro. With respect to the enhancement of amino acid incorporation, however, the site(s) of insulin action in this gland have not been studied. The studies proposed here, therefore, will be undertaken in order to examine the effects of streptozotocin diabetes and insulin in vitro on several cellular processes involved in the regulation of protein synthesis, including the membrane transport of amino acids and glucose, RNA synthesis, aminoacylation of tRNA, and the activity of the ribosome. In addition, calcium is postulated to play a role in mediating insulin action, and preliminary data suggest that the response of the rat submandibular gland to insulin stimulation is dependent upon the extracellular concentration of calcium. Therefore, the effects of varying calcium, both intracellular and extracellular, on the response of the submandibular gland will be examined. Although previous studies of insulin and the rat submandibular gland have been carried out using tissue slices, that particular experimental approach has been criticized. The studies proposed here, therefore, will be accomplished using dispersed submandibular cell aggregates in order to minimalize the effects of nonparenchymal cells that may impose diffusion barriers, and pericellular compartments which can alter the response of the cells to agonists. The long-term objectives of this research are (1) to examine the role of insulin in regulating submandibular gland metabolism, (2) to determine the site(s) of insulin action responsible for the enhancement of amino acid incorporation, (3) to study the role of calcium in modulating the response to insulin stimulation, and (4) to develop an exocrine cell model for the study of the mechanism(s) of insulin action. Despite the importance of insulin in regulating mammalian metabolism, the mechanism of insulin action has not been elucidated. It is hoped that in addition to increasing our knowledge of submandibular gland physiology, the results of these studies will provide an additional model for investigating the mechanism of insulin action at the molecular level.